The signaling pathways leading to the activation of the Rel/NFkappaB family of transcription factors are subjected to a novel level of regulation in which the ubiquitously expressed Rel/NFkappaB proteins are sequestered in the cytoplasm by inhibitor IkappaB proteins. The focus of the following research is to understand the molecular detains underlying the inhibitory mechanism employed by IkappaB proteins. The proposed research will provide a molecular for the behavior of the NFkappaB/IkappaB signaling pathway. We are using two different approaches for our investigations. High resolution X-ray crystallography will be integrated with biochemical and mutational analysis. Our research will address two central questions: First, how do IkappaB proteins selectively inhibit a unique subset of Rel/NFkappaB dimers. The proposed work relies on our ability to produce large amounts of highly pure proteins. This work has led us to obtain well-ordered crystals for structure analysis. Upon completion of the proposed experiments, the chemical rules that determine specificity and affinity of different NFkappaB/IkappaB complexes will be elucidated. These experiments will allow us to propose a model for how different complex formation between Rel/NFkappaB and IkappaB ultimately dictates the fate of a specific cell type.